Caspase-1, a cysteine protease, is critical for the production of mature IL-1&#946;and IL-18, two pro-inflammatory cytokines that play an important role in host defense, sepsis, and the pathogenesis of several inflammatory diseases. In addition, dysregulated IL-1&#946; production is major factor in the development of autoinflammatory disorders including familial cold autoinflammatory syndrome, Muckle-Wells syndrome, and neonatal-onset multiple-system inflammatory disease. These inherited autoinflammatory syndromes are caused by missense mutations in NLRP3, a member of the NOD-like receptor (NLR) family. NLR family members including NOD1, NOD2, NLRC4 and NLRP3 are intracellular proteins that are involved in the recognition of microbial components and activation of inflammatory pathways against invading pathogens. Recent work from several laboratories including our own indicate that NLRP3 is critical for the activation of caspase-1 through its interaction with the adaptor molecule ASC and the formation of the inflammasome, a molecular platform that drives caspase-1 activation. We have obtained evidence that NLRP3 plays an important role in the activation of caspase-1 and IL-1&#946;secretion in response to Toll-like receptor (TLR) ligands as well as infection with Staphylococcus aureus and Streptococcus pyogenes, two significant human pathogens. In addition, NLRP3 is critical for the activation of caspase-1 in response to particulate matter including silica and urate crystals. However, the molecular mechanism by which microbial and endogenous stimuli trigger caspase-1 activation via NLRP3 remains poorly understood. Our preliminary results revealed that TLR ligands and certain cytokines including TNF-&#945;, IL-1&#945;and IL-1&#946;promote NLRP3-dependent caspase-1 activation, at least in part, via NF-&#954;B activation. Unlike TLR agonists, we found that S. aureus and S. pyogenes activate the NLRP3 inflammasome via pore-forming toxins but independently of MyD88/TRIF and the purinergic P2X7 receptor. The goal of this proposal is to provide a better understanding of the molecular mechanism governing NLRP3 activation and the role of the NLRP3 inflammasome in host defense. Biochemical, genetic, and cellular approaches will be employed to study the function and activation of NLRP3. Given the important role of IL-1&#946;in immunity and inflammatory disease, understanding of the mechanism involved in caspase-1 activation and IL-1&#946;production via NLRP3 is expected to have a significant impact in the medical field